IRLF 


C   2 


THE  SPERMATOGENESIS  OF  LEPISMA  DOMESTICA 


BY 

HARRY  H.  CHARLTON 

(Osborn  Zoological  Laboratory,  Yale  University} 


DIS;  ION 


A  DISSERTATION 

presented  to  the  Faculty  of  the  Graduate  School  of  Yale  University, 
in  candidacy  for  the  Degree  of  Doctor  of  Philosophy 


1920 


Reprinted  from  JOURNAL  OF  MORPHOLOGY,  Vol.  35,  No.  2,  June,  1921 


AUTHOR'S  ABSTRACT  OF  THIS  PAPER  ISSUED  Reprinted  from  JOURNAL  OP  MORPHOLOGY 

BT  THE   BIBLIOGRAPHIC  SERVICE,    MARCH  28  Vol.  35,  No.  2,  June,   1921 


THE  SPERMATOGENESIS  OF  LEPISMA  DOMESTIC^ 

HARRY  H.  CHARLTON 

Osborn  Zoological  Laboratory,  Yale  University 

six  PLATES  (NINETY-FIVE  FIGURES) 

CONTENTS 

Introduction 381 

Material  and  methods 382 

Observations 384 

External  sexual  characters 384 

Male  reproductive  system 384 

Spermatogonia 384 

The  growth  period 386 

The  first  spermatocyte 389 

Resting  stage  of  the  second  spermatocyte 390 

The  second  maturation  division 391 

The  centrosome  in  the  spermatogonial  and  maturation  divisions 392 

The  spermatid 393 

The  spermatozoa 398 

Mitochondria 401 

Discussion 401 

Resting  stage  of  the  second  spermatocyte. 401 

The  idiochromosomes 402 

Synapsis  and  reduction 403 

The  centrosome 404 

The  acrosome 405 

The  middle-piece 406 

Comparison  with  Orthoptera 406 

Summary 408 

Bibliography 409 

INTRODUCTION 

Cytologists  have  long  found  the  class  Insecta  to  be  a  very 
fertile  field  for  investigation,  but  the  first  order  and,  according 
to  many,  the  most  primitive,  namely,  the  Thysanura,  seems  to 
have  been  neglected.  According  to  Harvey  ('16),  only  three 
papers  dealing  in  any  way  with  the  cytology  of  any  closely 

381 


382  HARRY   H.    CHARLTON 

related  forms  have  been  published.  The  writers,  Claypole  ('98), 
on  Anurida  maritima,  Lecaillon  ('01),  on  Orchesella  villosa,  and 
Willem  ('00),  on  Podura  aquatica,  simply  report  isolated  obser- 
vations which  are  necessarily  incomplete  and  limited  to  the  class 
Collembola. 

The  Lepismatoidea  have  therefore  never  been  made  the  sub- 
ject of  a  cytological  study,  and  it  was  in  the  hope  that  a  survey 
of  this  primitive  form  would  throw  some  light  on  the  present-day 
cytological  problems  that  this  investigation  was  undertaken. 

The  completion  of  the  study  shows  that,  instead  of  the  expected 
simplicity,  the  process  actually  is  a  complicated  one,  differing 
only  here  and  there  from  that  already  described  in  other  forms. 
These  differences,  however,  are  interesting  and,  together  with 
the  fact  that  it  is  the  first  cytological  work  in  a  new  class  of 
insects,  warrant  its  presentation. 

The  work  was  done  at  the  Osborn  Zoological  Laboratory  at 
the  suggestion  of  Professor  Petrunkevitch  beginning  in  the  fall 
of  1916.  During  1917-19  it  was  practically  suspended  except 
for  an  occasional  day  or  so  at  Columbia  University.  It  gives 
me  pleasure  to  express  my  thanks  to  Prof.  E.  B.  Wilson  for  his 
kindness  in  giving  me  laboratory  privileges  at  Columbia  and  to 
Prof.  Frank  R.  Lillie  for  facilities  accorded  at  the  Marine  Bio- 
logical Laboratory,  Woods  Hole,  Massachusetts,  during  the 
summer  of  1919. 

Most  of  all,  I  am  indebted  to  Prof.  Alexander  Petrunkevitch, 
first,  for  suggesting  the  problem  and  later  for  his  unfailing  help 
and  criticism. 

MATERIAL  AND  METHODS 

Lepisma  domestica,  commonly  called  the  fire-brat,  from  its 
frequently  observed  habit  of  running  apparently  unharmed  over 
hot  stones  in  bakeries,  belongs  to  the  class  Thysanura  of  the 
order  Lepismatoidea.  It  is  a  fairly  common  insect  in  New 
Haven,  and  can  be  kept  alive  in  the  laboratory  for  a  considerable 
period.  My  method  has  been  to  keep  them  in  large  glass  Stender 
dishes  without  covers,  since  the  insects  cannot  climb  up  a  clean 
glass  wall,  and  to  provide  them  with  a  cereal  such  as  corn  flakes 


SPERMATOGENESIS    OF   LEPISMA   DOMESTICA  383 

to  eat.  For  moisture  I  have  kept  shallow  dishes  filled  with 
moist  filter-paper  in  the  Stenders.  In  spite  of  these  precautions, 
in  the  course  of  a  month  or  so  the  creatures  begin  to  show  a 
shrinkage  of  the  abdominal  region  and  soon  die  with  the  posterior 
region  shrunken  fully  one-half. 

The  insects  were  either  killed  in  xylol  or  decapitated  and  the 
testes  dissected  out  immediately  in  physiological  salt  solution. 
As  soon  as  the  body  cavity  was  opened,  some  fixing  fluid  was 
introduced  by  means  of  a  fine  pipette.  This  renders  the  tissues 
more  opaque  and  makes  it  easier  to  locate  the  gonads  which  are 
loosely  surrounded  by  fat,  as  well  as  to  cause  better  fixation. 

For  counting  chromosomes,  Bouin's  fluid  at  38°C.  proved  the 
best,  but  for  general  fixation  of  the  cytoplasm  as  well  as  of 
chromatic  structures  nothing  equaled  Flemming's  strong  solu- 
tion. In  addition,  Hermann's  fluid,  Benda's  Flemming,  10 
per  cent  formalin,  Allen's  modification  of  Bouin,  Petrunkevitch's 
fluid,  and  Kopsch  were  tried  out  and  had  their  special  uses. 

The  testes  dissected  out,  fixed,  washed,  and  dehydrated,  were 
imbedded  in  paraffin  and  cut  into  sections  from  3  to  12  n  thick. 
Sections  of  7  p  thickness  were  found  to  be  very  satisfactory  for 
study,  and  in  general  this  was  the  thickness  used. 

The  stain  used  generally  throughout  was  Heidenhain's  iron 
haematoxylin  without  any  counterstain.  In  addition,  various 
counter  stains  were  tried,  and  also,  in  an  effort  to  get  a  selective 
stain  for  mitochondria,  Benda's  alizarin-crystal  violet  method. 
A  modification  of  CajaFs  silver-impregnation  method  by  Hortega 
(J16),  especially  recommended  for  centrosomes,  was  given  an 
inadequate  trial  with  but  fair  results. 

I  have  also  examined  a  number  of  splended  slides  stained  by 
the  safranin-gentian-violet-orange  G  method,  for  which  I  am 
indebted  to  Dr.  P.  W.  Whiting. 


384  HARRY   H.    CHARLTON 

OBSERVATIONS 
External  sexual  characters 

During  the  spring  months  of  March,  April,  and  May,  the  adult 
insects  are  in  the  best  condition  for  study,  but  since  it  takes 
some  time,  probably  a  year  or  more,  to  attain  sexual  maturity, 
the  early  stages  may  be  studied  at  any  time  of  year  in  young 
individuals. 

It  is  comparatively  easy  to  recognize  the  sexes  by  their  external 
appearance.  The  female  (fig.  2)  has  a  long  median  ovipositor 
extending  posteriorly  which  is  quite  prominent  in  the  living 
insect.  There  is  nothing  comparable  to  it  in  the  male  (fig.  1), 
for  the  penis  which  could  not  possibly  be  confused  with  the 
ovipositor  is  more  often  retracted  and  not  in  view. 

Male  reproductive  system 

The  testes  (fig.  3),  of  which  there  are  three  pairs  on  each  side 
of  the  middorsal  line,  occupy,  in  mature  individuals,  a  consider- 
able portion  of  the  anterior  two-thirds  of  the  abdomen.  The 
testes  lie  parallel  to  each  other,  extending  hi  a  ventroposterior 
direction,  and  each  is  connected  by  a  short  duct  with  the  vas 
deferens,  which  passes  as  a  straight  duct  posteriorly  where  it 
enlarges  to  form  the  seminal  vesicle.  From  the  seminal  vesicle 
a  similar  duct  extends,  which  soon  enlarges  considerably  and, 
after  bending  upon  itself  a  couple  of  times,  opens  into  the  base 
of  the  penis. 

Spermatogonia 

I  have  made  a  long  and  careful  search  for  primary  spermato- 
gonia  in  the  youngest  material  at  my  disposal,  which  consisted 
of  insects  only  2  or  3  mm.  long,  but  have  not  been  able  definitely 
to  identify  them.  It  is  therefore  very  probable  that  the  primary 
spermatogonia  occur  only  very  early  in  the  life-history.  Mun- 
son  ('06)  has  described  an  apical  cell  which  produces  early 
spermatogonia,  but  this,  too,  if  present  at  all,  would  be  found  in 
exceedingly  young  individuals. 


SPERMATOGENESIS    OF   LEPISMA   DOMESTICA  385 

One  does  find  cells  in  quite  large  numbers  at  the  blind  end  of 
the  testis,  which  differ  from  the  ordinary  spermatogonia  in 
having  large  homogeneous  nuclei  with  the  chromatin  condensed 
into  a  single  dense  mass  and  irregular  in  shape  (fig.  28).  I 
believe  these  to  be  immature  Sertoli  or  nurse  cells,  for^ater  on 
one  finds  such  cells,  only  now  they  are  larger,  more  elongated, 
and  contain  two  to  four  chromatic  bodies.  Those  at  the  region 
of  the  mature  spermatozoa  are  much  paler  in  color  and  may  be 
wrinkled  and  twisted  upon  themselves,  indicating  perhaps 
degeneration. 

The  spermatogonia  occupy  a  considerable  part  of  the  blind 
end  of  a  tubule  of  the  mature  insect  during  the  winter  and  early 
spring,  and  can  be  easily  recognized  by  their  position  and  by  the 
arrangement  of  the  chromatin  in  the  form  of  clumps  attached 
to  each  other  by  linin  threads  and  grouped  around  the  periphery 
of  the  nucleus.  This  arrangement  (fig.  4  is  a  surface  view)  is 
the  most  common  and  probably  represents  a  resting  condition. 
Although  I  have  not  been  able  to  count  these  clumps  of  chro- 
matin, the  number  is  easily  seen  to  be  more  than  the  haploid, 
and  each  one  probably  represents  a  spermatogonial  chromosome. 

The  nucleus  of  the  early  spermatogonium  is  quite  large  and 
almost  equal  in  size  to  the  nucleus  of  the  growth  period.  The 
two  or  three  spermatogonial  divisions  reduce  the  nuclear  and 
cell  size  by  apparently  not  allowing  tune  for  growth  between 
divisions  (figs.  8  to  11).  In  prophase  the  chromosomes  are  long 
and  bent  upon  themselves  and.  irregularly  scattered  throughout 
the  nucleus;  later  they  are  drawn  into  the  metaphase  plate  as 
shown  in  figure  5. 

It  is  only  in  the  larger  and  therefore  the  earlier  spermatogonia 
that  good  counts  of  the  chromosomes  can  be  made.  Figure  6 
shows  thirty-four  chromosomes  in  a  very  clear  metaphase  plate. 
The  chromosomes  are  of  the  curved-rod  type,  differing  con- 
siderably in  size,  but  close  observation  fails  to  show  any 
chromosome  or  group  of  chromosomes  behaving  in  any  way 
differently  from  its  neighbors. 

In  the  telophase  of  the  spermatogonial  division  the  chromatin 
becomes  granular  and  forms  a  more  or  less  eccentric  ring  around 


386  HARRY   H.    CHARLTON 

the  nuclear  wall  (figs.  11  and  16).  At  a  little  later  stage  one 
commonly  finds  an  irregular  clump  of  chromatic  material  repre- 
senting apparently  two  spermatogonial  chromosomes  lying 
against  the  nuclear  membrane  and  retaining  the  haematoxylin 
stain  (figs.  13,  14,  and  15).  For  these  chromosomes  I  shall  use 
the  term  idiochromosomes,  the  name  given  by  Wilson  ('05) 
and  meaning  'peculiar  or  distinctive  chromosomes.7 

In  some  presumably  young  cysts  the  spermatogonial  cells 
are  arranged  in  the  form  of  a  rosette,  their  median  ends  tapering 
toward  a  faintly  marked  open  center  and  showing  an  archo- 
plasmic  mass  or  sphere  (figs.  4  and  18).  Hegner  ('14),  Meves 
('97),  and  Shaffer  ('17),  as  well  as  others,  have  described  similar 
structures  and  considered  them  spindle  remains.  In  some  cases 
they  figure  them  as  extending  from  cell  to  cell.  While  this  is 
true  immediately  after  division  when  the  spindle  remains  are 
very  definite  (fig.  7),  it  is  not  possible  later  to  see  any  continu- 
ation or  connection  with  similar  bodies  in  adjacent  cells. 

In  the  spermatogonial  region  isolated  cells  are  occasionally 
seen  in  division,  but,  strangely  enough,  the  chromosomes  are 
paired  and  look  somewhat  like  tetrads  (fig.  32).  The  cells 
themselves  are  much  larger  than  the  spermatogonia  and  contain 
but  little  cytoplasmic  staining  material  in  the  form  of  a  flaky 
mass  at  either  end  of  the  cell  in  which  a  dark-staining  granule 
may  be  seen.  If  these  represent  division  in  the  Sertoli  cells, 
they  are  a  very  rare  occurrence.  In  the  older  Sertoli  cells  I 
have  occasionally  seen  evidence  indicating  division  by  amitosis. 

The  growth  period 

The  stages  of  the  growth  period  correspond  fairly  closely  with 
the  stages  described  by  Wilson  ('12).  After  the  telophase, 
chromosomes  of  the  last  spermatogonial  division  break  up  and 
form  a  granular  ring  just  inside  the  nuclear  wall,  the  chromatin 
arranges  itself  as  previously  described  in  the  form  of  clumps 
located  on  the  nuclear  membrane  (Wilson's  stage  b,  similar  to 
fig.  13).  In  heavily  destained  material  two  of  these  are  closely 
related,  one  of  them  being  flattened  against  the  nuclear  mem- 


SPERMATOGENESIS    OF   LEPISMA   DOMESTICA  387 

brane  and  retaining  the  dark  stain  of  the  haematoxylin  (fig.  15). 
In  addition  to  the  idiochromosomes,  a  similarly  staining,  small, 
spherical  granule  appears  (fig.  13).  The  chromatin  clumps  now 
become  granular  and  form  an  eccentric  'circle  against  the  nuclear 
membrane,  leaving  an  open  center  very  much  like  the  condition 
following  the  last  division.  The  homogeneous  granular  border 
is  at  first  deeply  stained,  but  later  loses  its  affinity  for  the  haema- 
toxylin and  appears  pale  in  color  (figs.  11  and  16). 

The  idiochromosomes  also  seem  to  break  up  into  unequal 
spherical  bodies,  three  to  eight  hi  number,  six  being  the  more 
common  number  (fig.  16) .  In  the  clear  central  region  the  remains 
of  the  preceding  spindle  are  quite  apparent.  Following  this 
stage  we  have  the  reappearance  of  the  idiochromosomes  (fig.  17), 
and  after  that  the  entire  nucleus  appears  granular,  the  central 
clear  area  disappearing  and  the  two  idiochromosomes  stand  out 
clearly  (fig.  21). 

It  has  not  been  possible  to  see  anything  like  an  unraveling 
stage  as  described  by  Wilson  ('12)  for  stage  c;  the  granular  con- 
dition being  directly  followed  by  delicate  threads  (Wilson, 
stage  d,  fig.  22)  which  seem  to  push  out  and  distort  or  break  the 
nuclear  wall.  This  is  soon  followed  by  the  synizesis  or  con- 
traction stage.  Here  the  threads  are  drawn  closely  together 
and  are  located  more  to  one  side  of  the  nucleus,  the  plasmosome 
and  idiochromosome  thread  often  remain  outside  of  the  con- 
tracted mass,  as  shown  in  figure  23. 

Popoff  ('08),  Gates  ('08),  and  Whiting  ('17)  look  upon  this 
as  due  to  a  rapid  absorption  of  water  by  the  nucleus;  in  other 
words,  an  osmotic  effect;  however,  it  has  often  been  considered 
an  artifact.  Although  at  this  stage  of  the  growth  period  the 
spireme  threads  stain  very  intensely,  making  it  difficult  to  trace 
the  individual  threads,  it  would  look  as  though  the  filaments 
became  arranged  in  the  form  of  loops  polarized  with  their  free 
ends  near  the  plasmosome  and  idiochromosome  threads.  Later 
on  when  the  threads  have  thickened,  this  bouquet  stage  is  much 
more  clearly  seen  (fig.  25) . 

It  has  not  been  possible  to  see  a  side-by-side  union  of  the 
spireme  threads,  the  synapsis  of  Moore  ('95),  but  the  number 


388  HARRY   H.    CHARLTON 

of  filaments  certainly  is  reduced  and  each  one  becomes  much 
thicker.  The  threads  now  loosen  up  and  occupy  practically 
all  the  cell,  the  space  between  the  nuclear  membrane  and  the 
cell  wall  being  quite  small  (fig.  24,  Wilson  ('12),  stage  f).  I 
have  not  been  able  to  find  the  longitudinal  splitting  of  the  thread 
—a  process  which  Wilson  ('12)  describes  as  taking  place. 

There  follows  a  period  when  it  is  hard  to  distinguish  the  threads 
as  such  (fig.  29,  Wilson  ('12),  stage  g).  Wilson  calls  it  a  net-like 
arrangement.  The  actual  breaking  of  the  threads  or  pachytene 
stage  is  not  well  exemplified  in  Lepisma,  but  stage  g  is  soon 
followed  by  the  clumping  of  the  chromatin  into  masses  irregular 
in  shape  and  joined  together  by  linin  threads  (fig.  30).  By  a 
further  condensation  of  these  masses  we  get  the  prochromosomes. 
The  formation  of  tetrads  showing  the  quadrivalent  condition 
of  the  autochromosomes  is  never  apparent,  neither  is  there  any 
split  indicating  a  parasynapsis. 

The  idiochromosomes  retain  their  form  and  staining  reaction 
until  the  formation  of  the  delicate  filaments  (stage  d),  when 
they  break  up  and  form  threads  which  are  darker  in  color  than 
the  other  threads,  and  one  may  be  seen  in  close  relation  to  a 
small  plasmosome  (fig.  35  a).  The  idiochromosome  threads  are 
at  first  very  long  and  may  extend  across  the  entire  width  of  the 
cell.  They  appear  somewhat  beaded,  just  as  is  the  case  with 
the  threads  of  the  autochromosomes. 

During  the  later  periods  the  threads  show  an  end-to-end  appo- 
sition, being  joined  by  very  fine  linin  fibers  (fig.  35  m).  The 
threads  now  become  shorter  and  thicker  assuming  the  U  shape 
followed  by  the  definite  formation  of  loops  with  the  plasmosome 
between  them  (figs.  20  and  35  j).  Figures  35  i  and  h  would 
seem  to  indicate  that  the  limbs  of  the  loop  come  together  and 
become  still  more  compact  to  form  clumps  lying  against  the 
nuclear  membrane  with  the  plasmosome  still  lying  between 
them.  For  a  considerable  time  the  idiochromosome  threads 
show  a  very  clear  inequality  in  that  the  thread  nearest  to  the 
plasmosome  is  the  longer  (fig.  35  k). 

A  second  small  plasmosome  may  be  formed  and  lies  to  one 
side  with  no  attachment  to  either  idiochromosome  thread  (fig. 


SPERMATOGENESIS    OF   LEPISMA   DOMESTICA  389 

35  e  and  f).  Later  I  believe  it  fuses  with  the  first,  for  the  latter 
is  seen  to  increase  considerably  in  size  and  to  show  at  times  a 
double  nature  (fig.  35  g  and  k). 

A  third  body  similar  in  shape  and  staining  reaction  to  that 
seen  in  the  spermatogonia  becomes  quite  prominent  at  this  time 
(figs.  29  and  30) ,  due  to  a  slight  increase  in  size  and  to  the  appear- 
ance of  a  clear  transparent  area  encircling  it.  Painter  ('14) 
describes  in  spiders  similar  small  dark-staining  spherical  bodies, 
which  he  calls  planosomes  and  which  first  make  their  appearance 
in  the  late  spireme  stage  and  which  he  was  able  to  trace  through 
the  succeeding  divisions.  The  planosomes,  according  to  him, 
have  spindle  fibers,  and  would  therefore  be  comparable  to 
chromosomes,  although  as  a  rule  they  do  not  divide,  but  linger 
near  the  middle  of  the  spindle  and  later  go  to  one  side. 

From  his  description  and  figures,  this  body  is  the  same  as  the 
one  found  in  Lepisma  domestica,  only  I  find  it  first  in  the  resting 
stages  of  the  spermatogonia,  and  have  not  been  able  to  follow  it 
beyond  the  prophase  stage  of  the  first  maturation  division. 

The  first  spermatocyte 

With  the  condensation  of  the  chromatin  segments  into  the 
prochromosomes,  the  nuclear  membrane  breaks  down  and  two 
chromosomes  located  near  the  periphery  are  seen  joined  together 
by  a  more  or  less  ribbon-like  connection,  forming  a  V-shaped 
structure.  Within  or  near  the  arms  of  the  V  the  plasmosome 
may  be  found  (figs.  36  and  37).  With  the  exception  of  the 
prophase  figures  in  which  the  idiochromosomes  stain  more  deeply, 
there  is  no  essential  difference  in  the  staining  reaction  of  the 
idiochromosomes  and  the  autochromosomes;  but  to  make  the 
behavior  of  the  idiochromosomes  plain  throughout  the  different 
stages  of  the  first  maturation  division,  they  have  been  drawn  in 
black,  while  only  the  outlines  of  the  autochromosomes  are  shown 
(figs.  36,  37,  41,  42,  43,  44,  and  45). 

The  chromosomes  arrange  themselves  on  the  spindle  and  in 
the  metaphase  plate  (figs.  38,  39,  and  40),  the  sex  or  idiochromo- 
somes are  still  connected  and  one  pair  of  the  chromosomes  is  a 


390  HARRY   H.    CHARLTON 

little  further  beyond  the  metaphase  plate,  so  that  in  plate  view 
one  pair  of  chromosomes  can  be  seen  to  be  at  a  different  level 
(figs.  39  and  40).  The  side  view  shows  how  one  limb  of  the  V 
extends  farther  than  the  other. 

The  metaphase  plate  (fig.  38),  in  which  the  idiochromosomes 
are  located  in  the  center  and  surrounded  by  a  ring  of  chromo- 
somes, reminds  one  of  the  arrangement  in  some  Hemiptera. 

There  is  little  change  in  the  position  of  these  joined  chromo- 
somes in  the  anaphase  (fig.  41),  except  for  a  shortening  of  the 
connecting  thread  and  possibly  a  slight  movement  of  the  whole 
toward  the  distal  pole.  Figures  42,  43,  and  46  picture  the 
telophase  arrangement,  the  idiochromosomes  going  undivided 
to  one  pole. 

There  are  sixteen  chromosomes  plus  the  two  idiochromosomes, 
or  eighteen  in  all,  in  the  first  spermatocyte  division.  Side  views 
have  not  been  counted,  owing  to  the  great  overlapping  of  the 
chromosomes.  The  plasmosome  may  be  identified  during  the 
late  prophase  (fig.  37),  but  not  definitely  after  the  actual  spindle 
formation.  Bodies  which  are  plainly  not  chromosomes  are 
often  seen  in  relation  to  the  spindle,  as  the  two  equal  bodies  in 
figure  40,  but  whether  these  represent  the  divided  plasmosome 
or  are  mitochondrial  is  not  conclusive. 

Resting  stage  of  second  spermatocyte 

In  the  telophase  of  the  first  or  early  prophase  of  the  second 
spermatocyte  (fig.  46),  the  chromosomes  are  breaking  up.  Some 
appear  unchanged,  while  others  have  swollen  to  a  spherical  shape 
and  stain  more  diffusely.  It  is  not  possible  to  identify  the 
idiochromosomes  at  this  time,  but  a  little  later,  when  the  resting 
nuclear  stage  is  reached,  the  double  nature  of  the  idiochromo- 
somes is  quite  apparent  as  the  nucleolus  in  one  of  the  now  divided 
cells  (figs.  47,  48,  50,  and  51).  It  is  not  possible  to  confuse  these 
resting  second  spermatocytes  with  the  early  spermatids,  because 
both  nuclear  and  cell  size  is  much  larger.  The  relative  sizes  of 
first  and  second  spermatocytes  and  spermatids  are  shown  in 
figure  33,  which  was  diagrammed  from  measurements  of  the  length 


SPERMATOGENESIS    OF   LEPISMA   DOMESTICA  391 

and  breadth  of  ten  representative  cells  of  each  kind,  and  the 
average  diameter  of  each  cell  and  of  the  ten  cells  taken. 

In  the  second  place,  the  chromatic  nucleolus  is  distinctly 
double  (fig.  47),  while  in  the  spermatid  it  is  single  and  smaller 
(figs.  64,  66,  and  68). 

During  the  growth  and  division  period,  spindle  remains  stand 
out  quite  clearly  as  one,  more  usually  as  two  vesicles,  formed 
probably  from  the  central  fibers  and  showing  a  granular  con- 
densation in  their  interior  (figs.  25  and  46). 

The  formation  of  the  resting  stage  and  the  subsequent  prophase 
is  a  rapid  one,  as  I  have  observed  resting  nuclei,  prophase,  and 
dividing  second  spermatocytes  in  the  same  cyst.  Figures  50 
and  51,  resting  and  prophase  stages,  respectively,  are  from  a 
slide  not  particularly  well  fixed,  as  the  cells  are  somewhat  swollen, 
but  figure  51  is  interesting  in  that  it  shows  the  formation  of 
spindle  fibers  before  the  nuclear  wall  has  broken  down  and  in 
figure  50  the  idiochromosomes  still  show  their  double  structure. 
The  resting  nucleus,  at  first  granular,  breaks  up  into  faintly 
staining  irregular  or  fantastically  shaped  entities  without  any 
visible  unraveling  stage  and  condense  quickly  into  the  pro- 
chromosomes  (figs.  49  and  50). 

The  second  maturation  division 

With  the  formation  of  the  spindle  for  the  second  maturation 
division,  two  types  of  metaphase  plates  are  seen:  one  (fig.  53) 
with  eighteen  chromosomes  and  another  (fig.  56)  with  sixteen. 
In  the  latter  case  I  have  one  perfect  anaphase  (fig.  59),  in  which 
both  plates  can  be  counted  and  both  show  sixteen  chromosomes. 

It  appears  that  the  idiochromosomes  are  now  equal  in  size 
and  no  longer  show  a  connecting  thread.  In  the  first  maturation 
division  the  idiochromosomes  were  distinctly  unequal,  but  each 
tapered  into  a  thread  connecting  it  with  the  other.  This  thread 
often  seemed  ribbon-like,  granular,  and  taking  the  iron  haema- 
toxylin  stain  like  the  chromosomes. 

It  seems  to  attain  its  maximum  length  at  the  metaphase  of 
the  first  maturation  division  and  to  shorten  a  great  deal  by  the 


392  HARRY   H.    CHARLTON 

time  the  telophase  is  reached,  and  it  would  appear  as  though 
this  thread  were  fused  with  the  smaller  idiochromosome  so 
that  they  both  appear  equal  in  the  metaphase  of  the  second 
maturation  division.  Another  factor  in  favor  of  this  hypothesis 
is  that  the  chromatic  nucleolus  of  the  resting  stage  shows  a  double 
structure  with  hardly  any  inequality. 

In  the  early  anaphase  (fig.  55)  all  the  chromosomes  show  a 
longitudinal  split  near  their  centers,  except  two  which  represent, 
I  believe,  the  divided  idiochromosomes.  The  anaphase  often 
shows  the  chromosomes  arranged  in  the  form  of  a  ring  (fig.  62). 
In  figure  63  the  chromosomes  are  at  the  poles  and  are  beginning 
to  form  a  nuclear  membrane,  but  no  change  has  taken  place  in 
the  centrosomes.  Figure  60,  a  late  telophase,  shows  that  one 
chromosome  differs  from  the  rest  in  being  elliptical,  while  the 
others  are  V-  or  U-shaped  and  slender.  A  still  later  telophase 
is  figured  in  figure  61,  the  chromatin  now  being  massed  at  the 
poles.  Two  types  of  spermatids  are  formed,  those  with  sixteen 
and  eighteen  chromosomes,  respectively. 

The  centrosome  in  the  spermatogonial  and  maturation  divisions 

In  the  archoplasmic  mass  or  sphere  representing  the  remains 
of  the  previous  spindle  one  may  occasionally  see  two  dark  gran- 
ules (fig.  18),  which  I  take  to  be  the  divided  centrosomes.  In 
the  division  figures  of  the  spermatogonia  centrosomes  are  difficult 
of  demonstration,  but  in  a  few  slides  I  can  make  them  out  as 
definite  single  granules  at  the  poles  of  the  spindle  (fig.  12).  I 
have  never  seen  astral  rays  or  anything  comparable  to  a  centro- 
sphere  at  the  time  of  division,  but  during  the  resting  stages  the 
centrosome  is  found  in  a  granular  sphere. 

From  the  division  figure  of  the  last  spermatogonial  mitosis 
until  shortly  before  the  synaptic  or  contraction  stage,  the  centro- 
some has  not  been  traced,  and  when  it  does  appear  a  considerable 
metamorphosis  has  taken  place.  At  about  the  time  when  the 
fine  spireme  threads  are  being  changed  into  loops,  a  granular 
mass  can  be  made  out  at  one  end  of  the  cell,  and  in  this  mass 
appear  two  short,  stubby  rods  lying  parallel  to  each  other. 


SPERMATOGENESIS    OF   LEPISMA   DOMESTICA  393 

Later  (figs.  20  and  26).  the  rods  lengthen  and  show  small  granules 
at  their  ends.  At  first  the  two  rods  form  an  angle  of  180°,  but 
this  angle  is  later  decreased  to  90°  or  less.  Each  rod  now  divides, 
but  the  halves  remain  attached  by  their  granule  ends^forming 
a  pair  of  V-shaped  centrosomes,  each  V  representing  a  divided 
centrosome.  This  whole  process  is  a  rapid  one,  for  all  stages  as 
well  as  the  separation  of  the  V7s  for  some  distance  may  be  seen 
in  cells  which  show  little  change  otherwise.  The  migration  is 
about  completed  and  the  V's  nearly  at  the  poles  by  the  time  the 
prophase  condition  is  reached  (fig.  30).  During  the  succeeding 
division  the  apex  of  the  V  is  directed  toward  the  chromosomes, 
while  its  limbs  touch  the  surface  of  the  cell.  The  V  may  open 
considerably,  nearly  to  a  straight  angle,  so  that  a  large  part 
of  the  outer  surface  of  the  rods  is  in  contact  with  the  cell  wall. 
The  cells  may  also  show  a  slight  depression  at  the  poles  (fig.  39) . 

The  spindle  fibers  all  lead  to  the  centrosome  region,  but  an 
actual  attachment  of  the  fibers  to  the  centrosomes,  while  taken 
for  granted,  does  not  show  clearly  in  sections. 

This  V  arrangement  can  be  identified  up  to  a  late  telophase  of 
the  primary  spermatocyte,  but  I  have  not  traced  it  through  the 
resting  stage  of  the  second  spermatocyte.  Each  second  sperma- 
tocyte would  receive  one  V,  but  when  the  rods  reappear  in  the 
division  figure  they  are  divided,  a  single  rod  at  either  pole  lying 
against  the  inner  surface  of  the  cell  wall  and  oriented  parallel 
to  each  other,  but  at  a  slight  angle  with  the  cell  axis.  The 
division  or  separation  of  the  V's  as  well  as  their  migration  to 
opposite  poles  must  take  place  during  the  resting  period. 

The  centrosome  rod  can  be  traced  through  every  succeeding 
stage  to  the  early  spermatid,  where  it  may  be  seen  lying  free 
in  the  cytoplasm  (fig.  65).  In  exceptional  cases,  as  in  figure  54, 
the  rods  have  granules  at  their  ends,  or  we  may  find  a  number  of 
granules  or  fragments  and  no  rod,  as  in  figure  58. 

The  spermatid 

The  young  spermatid  cell  is  considerably  smaller  than  the 
resting  stage  of  the  second  spermatocyte.  The  chromosomes 
clump  together,  form  a  nuclear  membrane,  and  quickly  break 


394  HARRY   H.    CHARLTON 

up.  The  nucleus  appears  round  in  polar  view,  but  oval  if  looked 
at  from  the  side.  Later  the  nucleus  becomes  spherical,  the 
chromatin  appearing  finely  granular  and  congregated  at  the 
boundaries  of  the  nucleus  leaving  an  open  center  (fig.  66).  One- 
half  the  cells  show  an  idiochromosome  nucleolus  which  usually 
presents  a  spherical  part  extending  into  the  nuclear  cavity  and 
a  flattened  area  against  the  inner  surface  of  the  nuclear  mem- 
brane, while  the  other  cells  do  not  possess  an  idiochromosome 
nucleolus. 

The  methods  of  fixation  and  staining  have  a  great  deal  to  do 
with  the  structures  observed  in  the  spermatid.  When  strong 
Flemming  is  used  for  fixing  followed  by  Heidenhain's  iron 
haematoxylin,  the  cytoplasm  of  the  early  spermatid  contains 
such  a  mass  of  intensely  staining  material  that  the  nuclear 
membrane  is  made  out  only  with  difficulty.  The  same  stain 
after  Bouin's  fluid  brings  out  the  nucleus  and  centrosomes,  but 
not  the  cell  inclusions. 

At  the  very  first,  the  cytoplasmic  structures  are  somewhat 
loosely  aggregated  around  the  nucleus,  but  particularly  between 
the  nucleus  and  the  last  division  plane,  The  centrosome  can 
easily  be  followed  from  the  telophase;  located  at  first  on  the  cell 
wall  of  the  dividing  second  spermatocyte,  it  later  moves  inward, 
occupying  the  space  between  the  cell  wall  and  the  nucleus 
(fig.  65). 

As  it  moves  around  to  get  between  the  nebenkern  and  the 
nucleus,  it  turns  90°  and  comes  to  lie  with  one  end  on  the  nuclear 
membrane  and  the  other  against  or  near  the  cell  wall  (figs.  66 
and  67).  The  rod-shaped  centrosome  now  frequently  shows  a 
granule  or  enlargement  at  the  nuclear  end. 

The  nebenkern  has  meanwhile  formed  a  broad  ring  of  densely 
staining  granular  material  in  the  center  of  which  spindle  remains 
of  the  last  division  appear  and  on  either  side  two  spherical  bodies 
become  visible  (fig.  69),  exactly  like  those  seen  in  the  two  matura- 
tion divisions,  and  undoubtedly  represent  old  spindles.  In 
cross-section  they  appear  as  rings  with  their  boundaries  staining 
in  varying  degrees,  often  looking  like  crescents,  and  may  possess 
a  darker  staining  center.  Looked  at  from  the  side,  they  take 
the  form  of  rods  with  faintly  stained  material  between  them. 


SPERMATOGENESIS    OF   LEPISMA   DOMESTICA  395 

When  the  centrosome  is  in  contact  with  the  nuclear  wall, 
one  usually  sees  a  granule  at  its  inner  end  (fig.  66),  and  later  a 
granule  similar  in  size  located  near  it  on  the  membrane  (fig.  67). 
This  suggests  the  breaking  away  or  division  of  the  granule  at 
the  base  of  the  centrosome. 

The  centrosome  may  now  change  its  position,  being  found  in 
the  region  of  the  nebenkern  or  even  at  the  opposite  side,  and 
shortly  the  delicate  axial  filament  is  seen  pushing  from  the 
cell  and  occasionally  carrying  a  small  clump  of  cytoplasm  with 
it,  very  much  as  has  been  described  by  Buder  (J15)  in  the  Lepi- 
doptera  and  called  by  him  'plasmaklumpchen.' 

The  single  granule  arising  from  the  centrosome  increases 
considerably  in  size  and  divides,  giving  rise  to  two  granules  which 
move  apart  and  come  to  lie  against  the  nuclear  membrane  and 
closely  applied  to  it  (figs.  65,  70,  71,  72,  and  73).  About  this 
time  or  a  little  later  a  somewhat  larger,  round  body  condenses 
out  of  the  nebenkern  ring,  as  shown  in  figure  72. 

Outside  of  the  breaking  up  of  the  idiochromosomes  and  a 
slight  tendency  to  become  pale  and  homogeneous,  the  nucleus 
remains  the  same  during  the  above  changes  in  the  cytoplasmic 
inclusions.  In  the  stage  which  follows,  the  delicate  axial  fila- 
ment is  quite  obvious  and  its  outgrowth  from  the  distal  end  of 
the  rod  centrosome  is  very  clear.  The  rod  has  swung  so  that 
now  it  is  in  contact  with  the  nucleus  throughout  its  entire  length 
and  the  thread  is  seen  traversing  the  space  between  nucleus  and 
cell  wall  (fig.  74).  The  nucleus  contains  numerous  dark-staining 
granules.  The  spindle  remains  are  prominent,  their  borders  have 
increased  in  thickness,  and  now  appear  as  irregular-shaped  thick- 
walled  vesicles.  The  nebenkern  ring,  cleared  of  the  spindle 
remains  and  of  the  various  granules  as  well  as  of  several  aggre- 
gations of  granular  mitochondria,  now  rounds  itself  up  into  an 
oval-shaped  dense  mass  which  later  becomes  round  (fig.  76). 
No  structure  is  at  first  apparent  except  a  heavily  stained  granular 
body,  but  one  soon  sees  a  vacuolization  of  its  border  and  we 
get  the  rosette  nebenkern  of  many  writers  (fig.  74) . 

Of  the  three  granules  already  mentioned,  those  arising  from 
the  centrosome  have  either  disappeared  or  have  become  so 


396  HARRY   H.    CHARLTON 

closely  adherent  to  the  nuclear  membrane  as  to  seem  a  part  of 
it,  while  on  the  other  hand  the  other  body  appears  slightly  larger 
(figs.  77  and  78). 

The  nucleus  continues  to  stain  darker,  due  to  the  enlargement 
of  the  chromatin  granules,  and  these  may  become  joined  to 
each  other  and  give  the  appearance  of  short  threads  (fig.  75). 
The  vacuolization  of  the  nebenkern  continues  at  the  expense  of 
the  central  body  which  becomes  smaller.  The  walls  of  the  pe- 
ripheral vacuoles  break  down,  the  spaces  becoming  larger  and 
larger,  until  there  is  but  one  vacuole,  which  may  exceed  even  the 
nucleus  in  size,  with  a  small  heavily  staining  central  part  (figs. 
80  and  81). 

From  this  period  on,  the  axial  filament  is  in  close  relation  to 
the  central  body  of  the  nebenkern,  which  in  well-fixed  material 
is  now  seen  to  be  made  up  of  a  spireme-like  thread.  I  have  been 
able  to  follow  it  throughout  the  greater  part  of  its  course  and  I 
feel  almost  certain  that  it  is  a  single  continuous  thread  (fig.  80). 
The  cell  now  begins  to  lengthen  somewhat  and  the  central  part 
of  the  nucleus  to  stain  heavily,  the  chromatin  moving  toward 
the  nuclear  center,  leaving  a  clear  transparent  border  (figs. 
78  to  83). 

Unless  one  is  fortunate  with  his  fixation  and  staining,  the 
central  part  of  the  nebenkern  shows  no  structure,  but  appears 
as  a  glassy  elliptical  body  suspended  in  the  single  large  vacuole 
by  means  of  the  axial  thread,  but  it  can  be  seen  very  clearly 
that  the  tail  filament  never  enters  the  central  body,  but  comes 
to  lie  against  it.  The  vacuole  membrane  lengthens  out  as  it 
increases  in  size,  while  the  central  thread-like  structure  breaks 
up  into  several  large  and  many  small  vesicles  (figs.  81  and  82). 
There  are  also  mitochondria-like  structures  located  between  the 
nebenkern  and  the  nucleus  as  well  as  some  distal  to  the  nebenkern. 

The  nebenkern  membrane  forms  apparently  the  sheath  of  the 
axial  thread,  some  cytoplasm  forming  clumps  around  the  distal 
part  of  the  thread,  but  the  vesicles  in  large  numbers  fill  the 
spaces  between  the  spermatozoa  as  they  increase  in  length. 
The  middle-piece  anlage  enlarges,  and  by  a  turning  of  the  nucleus, 
the  axial  filament  comes  to  lie  against  it  (figs.  85,  86,  88,  and  90). 


SPERMATOGENESIS    OF    LEPISMA    DOMESTICA  397 

The  body  then  flattens  out  against  the  nucleus  and  later  elon- 
gates slightly  (figs.  85  and  90).  The  nucleus  lengthens,  and  as 
it  does,  the  axial  filament  between  the  middlepiece  and  the 
centrosome  does  likewise.  The  centrosome,  however,  is  now  at 
the  apex  of  the  nucleus  and  will  hereafter  be  considered  ~as  the 
acrosome. 

At  the  time  when  the  nebenkern  membrane  and  its  vesicles 
have  completely  broken  up  and  are  only  apparent  as  end  products 
ensheathing  the  elongated  tails  or  located  between  the  filaments, 
the  nucleus  is  still  spherical,  compact,  and  does  not  take  the 
haematoxylin  stain  very  well  (fig.  82).  It  still  has  a  clear  area 
about  it  and  some  mitochondrial  material  about  the  middle- 
piece  anlage,  which  is  a  quite  prominent  body  located  usually 
on  the  opposite  side  of  the  nucleus  from  the  acrosome.  The 
axial  filament  arising  from  the  acrosome  at  the  apex  of  the 
nucleus  is  bent  backward  and  passes  near  the  middle-piece 
anlage. 

The  further  changes  are  the  loss  of  the  clear  ring  about  the 
nuclear  chromatin  by  the  spreading  out  of  the  chromatic  material. 
The  nucleus  shows  better  staining  qualities.  The  axial  filament 
comes  to  lie  nearer  to  the  middle-piece  and  the  latter  may  some- 
times show  one  or  more  bubbles  or  vesicles,  which  are  possibly 
mitochondrial,  in  relation  to  it  (figs.  84,  85,  and  86). 

The  nucleus  now  begins  slowly  to  elongate  and  seems  some- 
times to  pull  away  from  the  acrosome,  so  that  part  of  the  latter 
body  may  project  beyond  the  nucleus.  The  nucleus  continues 
to  lengthen,  the  chromatin  to  appear  paler  in  color.  The  mid- 
dle-piece enlarges  and  elongates  (figs.  88,  89,  90,  and  91).  The 
axial  filament  strand  between  the  acrosome  and  the  middle-piece 
remains  applied  against  the  nucleus  and  sometimes  may  show 
one  or  several  splits  in  the  thread,  leaving  an  elliptical  opening. 
In  material  stained  for  mitochondria,  a  cloud  of  granules  seems 
to  gather  about  the  thread  (fig.  87).  Although  it  cannot  be 
traced  directly,  I  am  of  the  opinion  that  this  axial  filament, 
which  is  loosely  applied  to  the  outer  nuclear  surface,  becomes  the 
undulating  membrane  of  the  mature  spermatozoon. 


398  HARRY  H.    CHARLTON 

Thompson  ('17,  p.  267)  suggests  the  formation  of  the  undulat- 
ing membrane  from  a  free  flagellum  in  the  Trypanosomes,  as 
follows:  "It  is  a  plausible  assumption  to  suppose  that,  as  the 
flagellum  waves  about  it  conies  to  lie  near  and  parallel  to  the 
body  of  the  cell,  and  that  the  frill  or  undulating  membrane  is 
formed  by  the  clear  fluid  protoplasm  of  the  surface  layer  spring- 
ing up  in  a  film  to  run  up  and  along  the  flagellum,  just  as  a  soap- 
film  would  be  formed  in  similar  circumstances."  Of  course  the 
axial  filament  in  this  case  is  located  between  the  nucleus  and  the 
outer  cell  wall,  but  it  seems  a  reasonable  hypothesis  to  think  of 
the  axial  filament  as  having  become  loose  from  the  nucleus 
and  as  able  to  draw  out  the  thin  layer  of  cytoplasm  some  little 
distance  from  the  nucleus  forming  the  undulating  membrane. 

The  nucleus  and  middle-piece  now  become  drawn  out  to 
considerable  length,  the  acrosome  decreases  in  size  and  we  see  a 
slight  projection  of  the  nucleus  extending  beyond  the  acrosome. 
The  elongated  nucleus  stains  darker  and  darker  until  no  structure 
can  be  made  out.  From  this  point  until  the  mature  spermatozoa 
are  reached  I  have  not  been  able  to  make  observations  (figs. 
93  and  94). 

The  spermatozoa 

A  study  of  the  mature  spermatozoa  has  been  made  by  teasing 
the  contents  of  the  seminal  vesicle  in  a  minute  quantity  of  physi- 
ological salt  solution  and  either  studying  them  alive  in  the  solu- 
tion or  fixing  the  teased  material  in  osmic  acid  fumes,  hot  corrosive 
sublimate,  Bouin  or  strong  Flemming,  and  staining. 

The  unstained  living  contents  make  an  interesting  study 
when  examined  by  means  of  the  dark-field  microscope.  In 
addition  to  the  spermatozoa  with  their  waves  of  movement 
extending  from  anterior  to  posterior  end  of  the  undulation 
membrane,  there  are  a  large  number  of  small  elliptical  bodies 
performing  active  brownian  movement.  I  thought  at  first  that 
these  bodies  were  the  true  spermatozoa  and  the  others  giant 
spermatozoa,  but  further  study  convinced  me  that  this  was  not 
the  case,  for  no  tails  could  be  found  upon  the  small  bodies, 
they  stained  only  by  plasma  stains,  and  furthermore  no  stages 
in  their  development  could  be  made  out. 


SPERMATOGENESIS    OF   LEPISMA   DOMESTICA  399 

Occasionally  the  lumen  of  the  vas  deferens  is  partially  filled 
with  granules  differing  in  size,  and  as  the  cells  lining  the  vas 
deferens  may  show  similar  granules  in  their  cytoplasm,  I  have 
considered  the  granular  material  of  the  lumen  to  be  secretion 
products  of  the  cells.  Although  the  bodies  present"  in  the 
seminal  vesicle  are  a  little  longer  than  broad  and  show  a  little 
difference  in  their  size  relations,  yet  I  think  they  represent  the 
secretion  found  in  the  vas  deferens.  Munson  ('06)  considers 
the  epithelial  cells  of  the  vas  deferens  of  the  butterfly  Papilio 
to  have  a  secretory  function,  but  unfortunately  he  does  not 
figure  or  describe  the  process. 

An  added  fact  of  interest  is  that  the  bodies  are  transmitted 
during  copulation  and  are  found  in  the  seminal  receptaculum 
of  the  female,  which  suggests  that  they  have  some  function  yet 
unknown  to  us. 

To  ascertain  their  nature,  smears  of  the  seminal  vesicle  were 
made,  fixed  in  hot  corrosive  sublimate,  and  stained  in  orcein, 
safranin  and  orange  G,  Delafield's  haematoxylin  and  orange  G, 
safranin  and  malachite  green,  safranin  and  bleu  de  Lyon,  Dela- 
field's  haematoxylin  and  erythrosin.  In  every  case  it  was  found 
that  the  granules  took  only  the  cytoplasmic  stains.  In  order  to 
test  the  possibility  of  the  granules'  being  of  a  fatty  nature,  fresh 
smears  were  stained  in  sudan  III,  but  the  result  was  negative. 
Smears  treated  with  ether  showed  also  no  effect  of  the  latter  on 
the  granules. 

When  the  spermatozoa  are  examined  in  the  fixed  and  stained 
condition  (text  fig.  A  and  fig.  95),  one  finds  a  long  chroma  tin 
staining  thread  ending  in  a  transparent  fine  point,  the  acrosome 
having  disappeared,  and  extending  from  near  the  apex  a  con- 
spicuous undulating  membrane.  The  free  edge  seems  formed  of 
a  little  denser  material  and  represents  in  all  probability  the 
proximal  part  of  the  axial  filament,  i.e.,  that  part  between  the 
acrosome  and  the  middle-piece. 

It  is  almost  impossible  to  see  just  where  this  membrane  leaves 
off  distally  as  it  gets  narrower  gradually,  but  I  should  say  that 
about  the  anterior  twro-thirds  of  the  spermatozoon  is  provided 
with  the  membrane.  It  is  not  possible  to  find  any  trace  of  the 


400  HARRY   H.    CHARLTON 

middle-piece  or  to  see  where  the  nucleus  leaves  off  and  the  tail 
filament  begins,  as  the  latter  structure  becomes  finer  and  more 
transparent  until  it  is  almost  impossible  to  see  where  it  ends. 


Text  fig.  A     Mature  spermatozoon  of  Lepisma  domestica  arranged  from  suc- 
cessive camera-lucida  drawings.      X  2800. 

I  have  noted  a  tendency  for  the  tails  of  the  living  spermatozoa 
to  stick  together.  The  spermatozoa  are  very  long,  measuring 
from  400  to  660/i. 


SPERMATOGENESIS    OF   LEPISMA   DOMESTICA  401 

Mitochondria 

Mitochondrial  structures  are  present  in  the  spermatogonia, 
but  in  very  small  numbers,  and  it  is  difficult  to  make  them  out. 
At  the  beginning  of  the  growth  period  they  appear  clearly  as  a 
dark-staining  crescent-shaped  mass  usually  located  at  one  end 
of  the  cell.  This  mass  soon  breaks  up  and  forms  some  six  to 
eight  bodies  differing  from  each  other  in  shape  and  size  (fig.  21), 
but  retaining  an  almost  constant  number.  These  bodies  take  the 
stain  intensely  and  appear  as  the  most  prominent  structures 
during  the  entire  growth  period.  As  the  cell  increases  in  size, 
the  mitochondria  becomes  so  conspicuous,  even  by  ordinary 
iron-haematoxylin  staining,  that  the  cytoplasm  seems  like  a 
dark  border  about  the  nucleus,  and  in  this  dark-staining  mito- 
chondrial  matrix  the  clumps  stand  out  clearly. 

During  the  first  maturation  division  the  ring  of  granular 
mitochondria  encircles  the  entire  spindle,  while  the  larger  bodies 
are  scattered  about  the  cytoplasm  and  are  located  near  but  not 
on  the  spindle.  The  mitochondrial  material  seems  to  divide 
equally,  half  the  granular  material  as  well  as  four  clumps  going 
to  each  cell  (fig.  39) .  In  the  resting  stage  of  the  second  sperma- 
tocyte  the  mitochondria  forms  a  narrow  dark-staining  ring 
about  the  nucleus,  but  the  clumps  are  no  longer  apparent  (figs. 
49,  50,  and  51).  In  figure  54  the  granular  mass  is  seen  arranged 
about  the  spindle  of  the  second  division,  while  figure  61  indicates 
how  they  gather  about  the  chromatin  at  the  poles  of  the  young 
spermatids.  In  the  description  of  the  spermatids  the  further 
history  of  the  mitochondria  has  already  been  given. 

DISCUSSION 

The  resting  stage 

While  in  many  animals  no  resting  nucleus  is  formed  following 
the  first  maturation  division,  the  chromosomes  of  the  telophase 
being  quickly  transformed  into  the  prophases  of  the  second 
division,  there  are  quite  a  number  of  exceptions  reported  in  the 
literature. 


•  •  «       .  • .  -  .     • 
-     .  • 


402  HARRY   H.    CHARLTON 

Murray  ('98)  finds  a  well-marked  resting  nucleus  in  the  Pul- 
monates,  Helix  and  Arion.  McGill  ('04)  found  it  to  happen 
occasionally  in  the  dragon-flies,  and  Painter  ('14)  describes  it 
as  occurring  in  the  spiders  with  the  accessory  chromosome 
persisting  as  a  nucleolus.  Kingsbury  ('01)  finds  in  the  salamander 
Desmognathus  fusca,  that  a  nuclear  membrane  is  formed  follow- 
ing the  first  maturation  division,  but  that  the  chromosomes 
never  lose  their  individuality. 

In  Lepisma  domestica  the  chromosomes,  with  the  exception 
of  the  idiochromosomes,  entirely  break  up  and  a  nuclear  mem- 
brane is  formed.  While  it  is  undoubtedly  of  short  duration,  still 
the  outward  individuality  of  the  autochromosomes  is  lost  and 
the  second  division  is  preceded  by  their  reformation. 

The  idiochromosomes 

Wilson  ('09)  divides  the  sexual  differences  of  the  chromosomes 
into  five  and  possibly  seven  types.  Lepisma  domestica  falls  hi 
line  with  his  type  IV  in  which  "the  male  has  a  pair  of  idiochro- 
mosomes, half  the  spermatozoa  receiving  both  and  hence  two 
more  than  the  other  half. " 

Only  one  form  has  been  found  which  has  this  arrangement, 
the  coreid  species  Syromastes  marginalis  L.  This  form  was 
first  described  by  Gross  ('04)  and  again  by  Wilson  ('09).  The 
accessory  chromosome  arises  by  a  synapsis  of  two  spermato- 
gonial  chromosomes  which  divide  equationally  in  the  first 
spennatocyte,  but  fail  to  divide  in  the  second. 

Lepisma  domestica  differs  in  that  the  two  spermatogonial 
chromosomes  do  not  fuse,  but  remain  separate  and  joined  by  a 
stout  thread.  They  pass  undivided  to  one  pole  in  the  first 
spennatocyte  division,  but  separate  in  the  second.  Wilson's 
prediction  that  the  female  of  Syromastes  would  have  two  more 
chromosomes  than  the  male,  he  afterward  found  to  be  the  case. 
Reasoning  in  a  similar  manner,  Lepisma  domestica  females  should 
have  thirty-six  chromosomes,  but  unfortunately  I  have  been 
unable  to  make  any  chromosome  counts  so  far  in  the  female. 


SPERMATOGENESIS    OF   LEPISMA   DOMESTICA  403 

Synapsis  and  reduction 

It  has  not  been  possible  in  the  ordinary  chromosomes  or  auto- 
chromosomes  to  see  whether  there  is  either  a  side-by-side  union 
of  the  spireme  threads,  a  parasynapsis,  or  an  end-to-end  "conju- 
gation, a  telosynapsis.  It  is  clear,  however,  that  the  spireme 
threads  in  postsynaptic  stages  are  much  thicker  and  are  present 
in  fewer  numbers.  Whether  they  are  half  the  leptotene  number 
or  not  could  not  be  made  out. 

In  the  case  of  the  idiochromosomes  the  conclusions  are  clearer. 
Each  idiochromosome  breaks  up  into  a  spireme  thread  and  the 
two  threads  eventually  unite  end  to  end,  one  of  them  being 
attached  to  a  large  plasmosome.  From  these  threads  two 
chromosomes  are  formed  by  the  condensation  of  the  chromatin, 
but  they  still  remain  united  by  a  thread  which  is  probably 
linin  in  nature  and  along  which,  when  the  thread  lengthens, 
the  chromatin  is  drawn  out. 

Synapsis,  or  a  side-by-side  conjugation,  if  it  takes  place  at 
all,  does  so  following  the  telophase  of  the  first  maturation  division. 
That  the  idiochromosomes  do  come  into  a  very  close  relation  is 
shown  by  the  longitudinal  split  apparent  in  the  idichromosome 
nucleolus  of  the  resting  nuclei  of  the  second  spermatocyte  (figs. 
47  and  50). 

If,  as  is  generally  conceded,  the  spermatogonial  chromosomes 
represent  two  groups,  one  of  maternal  and  the  other  of  paternal 
chromosomes,  and  the  homologous  pairs  conjugate  at  synapsis 
then  each  of  the  idiochromosomes  represents  one  spermatogonial 
chromosome.  A  further  proof  of  this  is  that  the  thirty-four 
spermatogonial  chromosomes,  judged  from  their  size,  are  all  of 
the  same  valence,  i.e.,  bivalent.  After  synapsis  the  autochro- 
mosomes  are  quadrivalent,  but  definite  four-part  tetrads  are  not 
apparent  during  the  prophase,  and  at  the  metaphase  the  chromo- 
somes are  dumb-bell-shaped,  the  longitudinal  pairing  leaving 
no  trace.  However,  in  one  cell  I  have  found  the  idiochromosomes 
at  the  time  of  anaphase  showing  a  bivalent  construction  (fig. 
45). 


404  HARRY   H.    CHARLTON 

The  first  maturation  division  separates  the  dumb-bell-shaped 
chromosomes  transversely  (fig.  39),  and  probably  represents  a 
reduction  division,  as  the  idiochromosomes  go  to  one  pole  undi- 
vided. The  second  division  of  the  autochromosomes  is  clearly 
a  longitudinal  one  (fig.  55),  while  the  idiochromosomes  separate 
transversely,  so  that  it  would  seem  that  this  represents  an 
equational  division  of  the  autochromosomes  and  the  separation 
of  the  idiochromosomes  one  from  the  other. 

The  centrosome 

The  single-  or  double-rod  type  of  centrosome  has  been  described 
in  a  variety  of  forms.  Meves  ('98)  and  Buder  ('15)  have 
described  them  in  the  Lepidoptera,  and  Sewertzoff  (Meves,  '00) 
in  Orthoptera,  and  Korff  ('01)  in  the  Coleoptera.  In  plants 
Von  Mottier  ('98)  found  them  in  the  tetraspore  mother  cell  of 
Dictyota  dichotoma.  Korff  also  found  them  in  the  sperm  cells 
of  the  domestic  hen  and  duck,  while  Hortega  ('16)  figures  them 
for  the  ganglion  cells  and  brain  of  man. 

Von  Mottier  does  not  consider  them  homogeneous,  but  to  arise 
from  small  granules.  In  the  beetles,  Korff  shows  them  to  be 
very  like  those  found  in  Lepisma  domestica,  but  he  has  not 
reported  any  granule  in  relation  to  the  rods  at  any  time.  He 
finds  the  limbs  of  the  V  separating  in  the  late  telophase  and 
appearing  parallel  to  the  polar  axis  in  the  spindle.  In  Lepisma 
domestica  the  centrosomes  are  always  oblique  to  the  axis,  but 
parallel  to  each  other. 

We  have  seen  the  centrosome  first  as  one  or  two  small  granules, 
then  as  double  rods  with  or  without  end  granules,  and  still  later 
as  single  rods  which  only  occasionally  show  a  granule.  In  very 
rare  cases,  instead  of  single  rods,  the  centrosome  consists  of 
several  granules  in  the  polar  position.  From  these  observations 
the  form  of  the  centrosome  would  seem  to  be  a  variable  quantity. 
It  is  interesting  in  this  regard  that  Korff  ('01)  was  only  able  to 
see  the  V-shaped  centrosome  in  the  sperm  cells  of  the  drake  and 
rooster,  while  all  the  other  cells  of  the  body  showed  centrosomes 
consisting  of  single  granules. 


SPERMATOGENESIS    OF    LEPISMA   DOMESTICA  405 

The  acrosome 

Although  a  number  of  the  older  writers,  notably  Platner  ('89), 
Niessing  (796),  Field  ('95),  and  Moore  (94'),  have  described  the 
acrosome  as  arising  from  the  centrosome,  Wilson  ('06)  comes  to 
the  conclusion  that  the  work  of  Henking  ('91),  Wilcox  ('96),  and 
Paulmier  ('99)  show  conclusively  that  in  the  insects  the  acro- 
some is  derived  from  the  nebenkern. 

That  this  is  not  the  case  in  Lepisma  domestica  can  be  easily 
proved,  for  in  every  stage  from  the  telophase  of  the  second 
division  until  the  oldest  transformation  stage  in  which  it  was 
possible  to  identify  structures,  the  centrosome  rod  and  its  change 
into  the  acrosome  can  be  followed. 

It  might  perhaps  be  argued  that  the  granule  is  really  the 
centrosome  and  the  rod  only  a  product  of  the  centrosome,  formed 
in  somewhat  the  same  manner  as  the  'battonet'  or  rodlet  in  the 
spermatid  of  the  Pribilof  fur  seal.  As  described  by  Oliver  ('13), 
it  arises  as  a  prolongation  from  the  anterior  centrosome.  In 
any  event,  the  acrosome  owes  its  origin  either  directly  or  indi- 
rectly to  the  centrosome. 

Goldsmith  ('19)  describes  in  the  tiger-beetle  a  condition,  which 
in  view  of  my  own  work  on  the  acrosome  and  middle-piece,  is 
very  suggestive.  He  figures  an  extra  nuclear  plate  or  middle- 
piece  which  is  formed  at  the  point  of  junction  of  the  axial  filament 
and  the  nucleus.  It  contains  several  chromatin-staining  bodies 
to  which  the  axial  filament  is  attached.  These  chromatin 
bodies  move  to  one  side  and  then  toward  the  anterior  end  of  the 
nucleus,  the  filament  coming  at  last  to  lie  against  the  elongated 
mitochondrial  body  (nebenkern)  and  the  bodies  to  assume  a 
bivalent  appearance  at  the  anterior  end  of  the  nucleus.  The 
middle-piece  becomes  drawn  out  into  a  granular  thread  con- 
tinuous with  the  axial  filament,  while  the  acrosome  appears 
later  and  fuses  with  the  other  two  bodies. 

It  would  appear  that  the  chromatin-staining  bodies,  to  which 
the  axial  filament  is  attached,  must  be  the  centrosomes,  and  that 
their  change  in  position,  due  to  the  rotation  of  the  nucleus,  is 
exactly  parallel  with  what  occurs  in  Lepisma  domestica. 


406  HARRY   H.    CHARLTON 

While  no  opinion  as  to  the  origin  of  the  acrosome  is  advanced, 
the  fact  that  it  arises  in  relation  to  the  granules  from  which  the 
axial  filament  originally  developed  would  point  to  the  probabil- 
ity of  a  centrosomal  origin.  The  granular  thread  which  he 
considers  to  be  the  drawn-out  middle-piece  is,  in  Lepisma 
domestica,  simply  the  proximal  end  of  the  axial  filament  carried 
to  the  apex  of  the  cell  along  with  the  centrosome,  and  it  would 
seem  as  though  such  an  interpretation  could  be  made  of 
Goldsmith's  results. 

The  middle-piece 

Wilson  ('06,  p.  337),  in  speaking  of  the  essential  structures 
in  a  spermatozoon,  lists  the  middle-piece  as  a  body  Which  "  either 
contains  a  formed  centrosome  or  a  pair  of  centrosomes,  or  is 
itself  a  metamorphosed  centrosome. " 

It  is  altogether  possible  that  the  middle-piece  in  Lepisma 
domestica  arises  from  one  or  both  of  the  granules  which  we 
found  to  have  their  origin  from  the  centrosome  and  later  traced 
them  to  where  they  were  closely  applied  to  the  nuclear  wall. 
In  fact,  we  would  naturally  expect  something  of  the  kind,  but 
unfortunately  the  later  history  of  the  granules  could  not  be 
followed.  The  question  is  further  confused  by  the  presence  of 
a  body  which  condenses  from  the  nebenkern  ring  and  which, 
from  its  size,  staining  power,  and  position,  would  point  to  its 
becoming  the  middle-piece.  Furthermore  this  body  can  be 
found  practically  in  all  the  stages  up  to  the  apposition  of  the 
middle-piece  to  the  nucleus,  but  here  again  we  are  stopped,  for 
we  have  not  seen  the  actual  formation  of  this  body  into  the 
middle-piece.  It  is  possible  that  a  further  study  will  solve  this 
difficulty. 

Comparison  with  Orthoptera 

At  the  first  glance  there  seems  to  be  a  similarity  between  the 
spermatogenesis  in  Thysanura  as  described  above  and  that  in 
Orthoptera  as  previously  described  and  particularly  given  by 
Payne  ('16)  for  Gryllotalpa  borealis,  but  there  are  also  differences 
which  cause  one  to  question  whether  the  resemblance  is  not 
more  superficial  than  real. 


SPERMATOGENESIS    OF   LEPISMA   DOMESTICA  407 

In  the  first  place,  the  general  shape  and  arrangement  of  the 
chromosomes  in  the  spermatogonia  of  Gryllotalpa  borealis  are 
very  much  like  similar  stages  in  Lepisma  domestica.  Payne 
also  has  described  changes  in  the  mitochondrial  mass  of  the 
spermatid  (his  figures  E,  F,  and  G,  pi.  2),  which  have  almost 
exact  counterparts  in  Lepisma  domestica.  Then  again  his 
figure  J  on  plate  3  shows  an  axial  filament  in  which  the  cytoplasm 
is  so  arranged  in  waves  as  to  look  like  the  undulating  membrane 
found  in  the  Thysanura. 

A  comparison  of  the  group  of  chromosomes  associated  prob- 
ably with  sex  in  the  two  forms  shows,  however,  several  important 
differences.  In  Gryllotalpa  borealis  Payne  finds  a  single  chromo- 
some which  does  not  divide  in  the  first  maturation  division  and 
therefore  could  be  directly  compared  with  the  idiochromosomes 
of  the  Thysanura  were  it  not  for  the  fact  that  the  single  chromo- 
some is  associated  with  an  unequal  bivalent  chromosome  and  in 
division  always  goes  to  the  same  pole  as  the  large  'end'  of  the 
unequal  chromosome.  Therefore,  the  two  resulting  secondary 
spermatocytes  differ  not  only  in  that  one  has  an  extra 
chromosome,  but  also  in  that  the  same  cell  possesses  the  large 
'end'  of  the  unequal  chromosome,  while  the  smaller  part  passes 
to  the  other  cell.  Payne  favors  the  view  that  these  chromosomes 
represent  a  triad  group  rather  than  an  unequal  pair  of  idiochro- 
mosomes and  an  accessory  chromosome. 

Payne  has  not  been  able  to  trace  the  centrosome  of  the  second 
maturation  division  through  to  the  spermatid,  and  in  fact  has 
not  been  able  to  demonstrate  the  presence  of  the  centrosome  in 
the  spermatid  at  all,  although  he  presumes  that  the  body  from 
which  the  axial  filament  arises  and  which  later  becomes  the 
middle-piece  may  be  a  centrosome.  Further,  he  describes  the 
acrosome  as  arising  from  an  elongated  body  which  suddenly 
appears  de  novo  in  the  cytoplasm,  whereas  in  Lepisma  domes- 
tica the  acrosome  is  formed  from  a  rod-like  centrosome. 


JOURNAL  OF  MORPHOLOGY,  VOL.  35,  NO.  2 


408  HARRY   H.    CHARLTON 

SUMMARY 

1.  The  male  Lepisma  domestica  has  three  pairs  of  testes  on 
each  side,  each  testis  connected  by  a  duct  with  the  respective 
vas  deferens. 

2.  The  blind  end  of  the  testis  contains  the  youngest  stages. 

3.  Primary  spermatogonia  are  formed  very  early  in  life. 

4.  Thirty-four  chromosomes  are  present  in  the  spermatogonia. 
A  chromatic  nucleolus  is  present  in  the  resting  stage  of  the 
spermatogonia. 

5.  The  growth  stages  follow  the  description  given  by  Wilson 
('12). 

6.  A  planosome  is  seen  in  the  resting  stage  of  the  sperma- 
togonia appearing  in  the  growth  stages  as  a  much  larger  body. 

7.  One  or  two  plasmosomes  appear  shortly  after  formation  of 
the  spireme  threads,  and  disappear  later. 

8.  There  are  eighteen  chromosomes  in  the  first  maturation 
division.     The   two    idiochromosomes   pass   undivided    to    one 
pole. 

9.  The  autochromosomes  divide  longitudinally  in  the  second 
division,    while   the   idiochromosomes   do   not.     Instead,    they 
separate,  each  spermatid  receiving  one  idiochromosome. 

10.  The  form  of  the  centrosome  is  changeable,  but  its  almost 
constant  presence  either  in  the  shape  of  a  granule  or  of  a  rod 
indicates  that  it  may  be  a  permanent  cell  structure. 

11.  A  chromatic  nucleolus  is  present  in  half  of  the  spermatids. 

12.  The  nebenkern  is  formed  from  granular  mitochondria,  the 
remains  of  the  last  and  of  two  previous  spindles. 

13.  The  axial  filament  grows  from  the  end  of  the  rod-shaped 
centrosome  which  forms  the  acrosome. 

14.  Another  body,  presumably  derived  from  the  nebenkern, 
forms  the  middle-piece. 

15.  The  nebenkern,  after  separating  out  the  spindle  remains 
and  several  accumulations  of  mitochondrial  material,   form   a 
vacuolated  body  which  furnishes  a  sheath  for  the  axial  filament. 

16.  The   axial  filament  persists   and  forms   the   undulating 
membrane  of  the  mature  spermatozoa. 


SPERMATOGENESIS    OF   LEPISMA   DOMESTICA  409 

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EXPLANATION  OF  PLATES 

All  the  figures,  with  the  exception  of  the  first  three,  were  made  with  a  Zeiss 
2-mm.  apochromat.  objective  and  Zeiss  compensating  ocular  no.  12.  In  order 
to  get  as  high  a  magnification  as  possible,  drawings  were  made  at  table  level,  giving 
an  enlargement  of  about  1850  diameters. 


ABBREVIATIONS 


a,  acrosome 
a/,  axial  filament 
c,  centrosome 
it  idiochromosomes 
m,  mitochondria 
mp,  middle-piece 
n,  nebenkern 
o,  ovipositor 


p,  penis 

pi,  plasmosome 

ps,  planosome 

s,  spindle  remains 

sv,  seminal  vesicle 

t,  tubules 

vd,  vas  deferens 

x,  middle-piece  anlage 


411 


PLATE  1 

EXPLANATION   OF  FIGURES 

1  Posterior  segments,  ventral  surface,  of  male  Lepisma  domestica.     X  20. 

2  Posterior  segments,  ventral  surface,  of  female  Lepisma  domestica.     X  20. 

3  Testes  of  one  side  showing  connection  with  vas  deferens  and  seminal  vesicle. 
X  20. 

4  Spermatogonium,  surface  view,  showing  resting  condition  of  nucleus  and 
the  attraction  sphere. 

5  Prophase  of  spermatogonium. 

6  Metaphase  plate  early  spermatogonium.    Thirty-four  chromosomes  joined 
by  linin  threads. 

7  Telophase  spermatogonia  showing  persistent  spindle  remains. 

8,  9,  10,  and  11    Nuclei  of  spermatogonia  decreasing  in  size  with  each  division. 
12    Spermatogonial  metaphase  from  the  side.    Centrosome  as  single  granule  at 
poles. 

13,  14,  and  15    Resting  stages  spermatogonia  to  show  idiochromosomes. 

16  Spermatogonium  showing  the  breaking  up  of  the  idiochromosomes. 

17  Beginning  of  growth  period.     Idiochromosomes  reformed. 

18  Spindle  remains  from  spermatogonium  with  two  centrosome  granules. 

19  Same  as  the  preceding  from  early  growth  period  with  V-shaped  centro- 
somes. 

20  Centrosome  rods  with  granules  at  their  ends,  before  division.    Plasmosome 
and  idiochromosome  loops. 


412 


SPERMATOGENESIS  OF  LEPISMA  DOMESTICA 

HARRY   H.   CHARLTON 


PLATE  1 


20 


PLATE  2 

EXPLANATION  OF   FIGURES 

21    First  spermatocyte.    Nucleus  homogeneous  except  for  idiochromosomes. 
[22    First  spermatocyte  showing  fine,  loosely  arranged  leptotene  thread.    Idio- 
chromosomes very  pale  in  color. 

23  Early  contraction  stage  of  first  spermatocyte.     Plasmosomes  with  idio- 
chromosome  thread  to  one  side. 

24  First  spermatocyte  showing  a  spreading  out  of  the  threads  following  con- 
traction.    Plasmosome  and  idiochromosome  threads  shown  in  surface  view. 

25  Bouquet  stage  of  first  spermatocyte.     Only  a  few  loops  shown  in  drawing. 

26  Pachytene  stage  of  first  spermatocyte. 

27  Same  as  the  preceding,  only  somewhat  later. 

28,  31,  and  34    Sertoli  cells  associated  with  different  stages  in  the  development 
of  the  spermatozoa. 

29  Confused  or  net-like  stage  of  first  spermatocyte.     Planosome  prominent. 

30  Early  prophase  of  first  spermatocyte.     Centrosomes  have  migrated  nearly 
to  poles.    Two  plasmosomes  present. 

32    Dividing  cell  from  spermatogonial  region.     Possibly  a  Sertoli  cell.     Chro- 
mosomes paired. 

tx*3    Diagram  to  show  the  relative  sizes  of  cells  of  first  and  second  maturation 
divisions  and  of  the  spermatid. 


414 


SPERMATOGENESIS  OF  LEPISMA  DOMESTICA 

HARRY   H.   CHARLTON 


PLATE  2 


34- 


415 


PLATE  3 

EXPLANATION   OF   FIGURES 

35  The  plasmosomes  and  idiochromosomes  during  the  growth  period  of  the 
first  spermatocyte  showing  formation  of  spireme  threads  and  their  contraction 
into  the  prochromosomes. 

36  and  37   Prophase  of  first  spermatocyte  showing  connected  idiochromosomes. 
38    Metaphase  plate  of  first  spermatocyte  with  idiochromosomes  in  center. 

c-39  Side  view  metaphase  of  first  spermatocyte  to  show  dumb-bell-shaped  chro- 
mosomes and  their  transverse  division.  Idiochromosomes  joined  together. 

40    Same  as  the  preceding. 

41,  42,  43,  and  44  Side  views  of  dividing  first  spermatocyte,  showing  idio- 
chromosomes during  anaphase  and  telophase. 

45  Oblique  view  of  anaphase  of  dividing  first  spermatocyte.  Thirty-three 
chromosomes  plus  the  idiochromosome  complex  which  here  shows  each  idiochro- 
mosome  as  bivalent. 


416 


SPERMATOGENESIS  OF  LEPISMA  DOMESTICA 

HAKBY  H.   CHARLTON 


PLATE  3 


PLATE  4 

EXPLANATION  OF  FIGURES 

46  Early  second  spermatocyte,  showing  the  breaking  up  of  the  telophase 
chromosomes.     Position  of  spindle  remains  typical. 

47  Resting  stage  of  second  spermatocyte. 

48  Same  as  the  preceding,  the  chromatic  nucleolus  found  in  only  half  the  cells 
of  the  second  spermatocyte. 

49  Early  prophase  of  second  spermatocyte. 

50  Same  as  the  preceding,  chromatic  nucleolus  double.    Ring  of  mitochondrial 
granules  around  the  nucleus. 

51  Resting  stage   of  second  spermatocyte,   showing  formation   of  spindle 
fibers  before  the  prophase  stage  is  reached. 

u  52    Side  view  metaphase  of  second  spermatocyte. 

53  Eighteen  chromosome  metaphase  plate  of  the  second  spermatocyte. 

54  Side  view  metaphase  plate  of  second  spermatocyte.     Rod  centrosomes 
with  granules  at  inner  ends. 

55  Early  anaphase  of  second  spermatocyte  with  the  autochromosomes  divid- 
ing longitudinally,  while  the  idiochromosomes  have  separated  transversely. 

56  Metaphase  plate  of  second  spermatocyte.     The  sixteen  chromosome  type. 

57  Later  anaphase  of  second  spermatocyte. 

58  Same  as  the  preceding  with  granules  at  poles  instead  of  rods. 


418 


SPERMATOGENESIS  OF  LEPISMA  DOMESTICA 

HAKRY   H.   CHARLTON 


PLATE  4 


419 


PLATE  5 

EXPLANATION   OF   FIGURES 

59  Two  anaphase  drawings  of  second  spermatocyte  from  same  cell.     Sixteen 
chromosomes  in  each. 

60  Spermatids  from  division  of  eighteen  chromosome  second  spermatocyte. 
The  divided  idiochromosomes  differ  in  shape  and  are  darker  stained  than  the 
autochromosomes. 

61  Young  spermatids  before  the  reorganization  of  nucleus. 

62  Anaphase  of  second  spermatocyte,  showing  ring-like  arrangement  of  chro- 
mosomes. 

1  63  Early  formation  of  nucleus  of  spermatid. 

64  Spermatid  with  resting  nucleus  and  nucleolus. 

65  Early  spermatid  with  rod  centrosome. 

66  Same  as  the  preceding,  showing  centrosome  with  granule  at  base  lying 
against  nuclear  membrane. 

67  Same  as  the  preceding.     The  granule  broken  off. 

68  Same  as  the  preceding.     The  granule  much  enlarged. 

69  Elements  of  nebenkern  from  early  spermatid. 

70  Spermatid  showing  division  of  granule. 

71,  72,  and  73    The  nebenkern  ring  from  spermatid,  showing  separation  of 
granules  and  formation  of  new  body  from  the  nebenkern. 

74  Central  part  of  nebenkern  of  spermatid,  showing  rosette  form  and  spindle 
remains. 

75  Spermatid  showing  vacuolization  of  nebenkern  border  further  advanced. 
Nuclear  granules  larger  and  stain  darker. 

76  Spermatid  showing  centrosome  applied  to  nuclear  membrane. 

77  and  78    Spermatid  showing  clear  space  forming  around   chromatin   of 
nucleus.    Middle-piece  anlage  present. 


420 


SPERMATOGENESIS  OF  LEPISMA  DOMESTICA 

HARRY   H.   CHARLTON 


PLATE  5 


61 


69 


72 


PLATE  6 

EXPLANATION   OF   FIGURES 

79  Same  stage  as  figure  78.     Centrosome  rod  shown  clearly. 

80  Spermatid   with    chromatin    of   nucleus   more    condensed.     Thread-like 
structure  in  middle  of  nebenkern  vacuole.    Mitochondria  between  nebenkern 
and  nucleus  as  well  as  distal  to  nebenkern. 

81  Same  as  the  preceding,  but  threads  of  nebenkern  have  broken  and  now 
form  vesicles. 

82  Same  as  the  preceding  showing  linear  arrangement  of  small  vesicles. 

83  Spermatid  of  a  slightly  later  state  than  the  preceding. 

84  Spermatid  with  nebenkern  contents  thrown  off. 

85  and  86    Spermatids.     Side  and  surface  view  adjacent  cells  to  show  relation 
of  axial  filament  to  middle-piece  anlage.     Filament  thicker  on  account  of  mito- 
chondrial  granules  in  relation  to  it. 

87  Spermatid  showing  mitochondrial  granular  mass  in  relation  to  middle- 
piece  and  axial  filament. 

88  Spermatid  showing  secondary  relationship  to  middle-piece  anlage. 

89  Spermatid.     A  later  stage  to  show  mitochondrial  ribbon  about  the  proxi- 
mal part  of  the  axial  filament. 

90  Spermatid  nucleus  elongating  and  staining  darker.     Middle-piece  getting 
longer. 

91  Same  as  the  preceding. 

92  Elongation  of  nucleus  of  spermatid.     Mitochondria  about  filament. 

93  Spermatid.     A  still  later  stage,  no  mitochondria  shown. 

94  Spermatid  nucleus  very  much  drawn  out.     Acrosome  at  apex  and  darker 
part  indicates  position  of  middle-piece. 

95  Part  of  anterior  end  of  mature  spermatozoon.     The  undulating  membrane 
with  its  thicker  border  is  well  shown. 


422 


SPERMATOGENESIS  OF  LEPISMA  DOMESTICA 

HARKY  H.  CHARLTON 


PLATE  6  • 


423 


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